Oxidative Phosphorylation part 2: Making ATP

electrons from cytoplasmic NADH are moved from the
cytoplasm to the matrix by

in muscle cells

the glycerol 3-phosphate shuttle

electrons from cytoplasmic NADH are moved from the
cytoplasm to the matrix by

heart/liver/other cells

the malate-aspartate shuttle

cytoplasmic glycerol 3-phosphate dehydrogenase (G3PDH)

What transfers electrons
from NADH to DHAP,
converting it to G3P

mitochondrial G3PDH

What converts G3P back to DHAP,
transferring the electrons to
FAD to make FADH2

FADH2 transfers electrons to
reduce Q to QH2, which
brings them to Complex what

III

other NADH molecules yield
2.5 ATP, but because these
cytoplasmic NADH electrons
bypass Complex I, they yield
how many ATP per NADH

1.5

In the heart and liver, electrons from cytoplasmic NADH are used to generate
mitochondrial NADH through the malate–aspartate shuttle, which uses how many
membrane transporters and how many enzymes

2, 4

Using the malate–aspartate shuttle, the mitochondrial NADH enters at Complex what (as usual) and yields 2.5 ATP

I

promotes symport of one H2PO4-
and one H+ into the matrix

phosphate translocase

antiporter that moves ADP into
the matrix and ATP out at the
same time

adenine nucleotide translocase

adenine nucleotide translocase
- makes up 15% of the protein
in the mitochondrial what
membrane

inner

adenine nucleotide translocase
- we exchange our body weight
in ATP per day (what
concentration in cytoplasm
than matrix)

higher

how many protons are used to bring
one phosphate in to make ATP

1

a complex of mitochondrial ATP synthase and
both translocases

ATP synthasome

In 1961, Peter Mitchell proposed the
chemiosmotic hypothesis which stated that
most ATP synthesis in respiring cells comes from
an electrochemical gradient of protons across the
inner membrane of mitochondria formed using
the energy from NADH and FADH2 acquired
during the what breakdown of energy-
rich molecules such as glucose.

oxidative

Mitochondrial ATP synthase

the large enzyme complex believed to make
ATP in the mitochondrial membrane was purified in 1966.

However, the lab that discovered it was unable to coax it into making ATP in
what (it was first called mitochondrial ATPase because it did the reverse
reaction)

vitro

bacteriorhodopsin

In 1974, Racker and Stoeckenius showed
that if protons were pumped inside an
artificial membrane using a proton pump
called what, then ATP synthase
could generate ATP with no intermediate
needed

chemiosmotic theory

electron
transport and ATP
synthesis are
coupled by a proton
gradient across the
inner mitochondrial
membrane created
by the electron
transport chain

proton-motive force

the energy stored in an
electrochemical proton
gradient across the
mitochondrial inner
membrane

potential

proton-motive force =
composed of chemical
and electrical what
energy

removal of H+ from the matrix
by the ETC generates:

what potential – the matrix
side becomes negative
compared to intermembrane
side (membrane potential of
0.14 V)

electric potential

removal of H+ from the matrix
by the ETC generates:

what potential – the matrix
side has a higher pH than the
intermembrane side (1.4 units
higher)

chemical potential

mitochondrial ATP synthase/F0F1 ATP synthase

a multiprotein complex that drives the synthesis of ATP as
protons flow passively back into the matrix through it

Mitochondrial ATP Synthase
contains two distinct components:

which: integral membrane protein that has channels through which
protons flow back into the matrix from the intermembrane side

F0

Mitochondrial ATP Synthase
contains two distinct components:

which = protrudes into the matrix and synthesizes ATP

F1

ATP synthases bind to one another to form what, which then
oligomerize. The oligomers contribute to the formation of cristae.

dimers

subunits

F0 subunit:
c ring composed of 8 -14 c
what that spans the inner
membrane
(number depends on species
– all vertebrates have 8)

F0 subunit:
one a subunit that binds the
what of the ring

outside

F1

F0 subunit:
a column made of b subunits
and d subunit that connects to
what subunit

F1 subunit:
how many a subunits and how many
b subunits in a hexameric ring

3, 3

F1 subunit:
each what subunit has an active
site for ATP synthesis

b

F0

F1 subunit:
a central stalk composed of
g and e subunits runs up the
middle of the ring and into the
what subunit in the membrane

Mitochondrial ATP Synthase
Each enzyme has how many active sites located on the three β
subunits

3

Mitochondrial ATP Synthase
Each β subunit can be in one of three states based on its
interaction with the which subunit stalk

γ

Which state - nucleotides enter or leave the β subunit

O (open, empty)

Which state - nucleotides are trapped in the β subunit

L (loose, ADP)

Which state - ATP is synthesized from ADP and Pi

T (tight, ATP )

conformations

Proton Flow Through ATP Synthase Leads to
the Release of Tightly Bound ATP
Each subunit cycles through the three what

shifts

Binding-Change Mechanism for ATP Synthase
The rotation of the γ subunit interconverts the β subunits
by causing conformation what

inding-Change Mechanism for ATP Synthase
The rotation of the γ subunit interconverts the β subunits
by causing conformation shifts
- rotation is what

counterclockwise

Rotational Catalysis: the World’s Smallest Molecular Motor
The rotation of the γ subunit was observed directly
movement of the γ subunit was visualized as a result of ATP
hydrolysis (what by itself can be an ATPase)

F1

Rotational Catalysis: the World’s Smallest Molecular Motor

The hydrolysis of a single ATP powered the rotation of the γ
subunit what° (single steps - what turns)

120

Binding-Change Mechanism for ATP Synthase
The rotation of the γ subunit interconverts the β subunits.
as the γ subunit rotates 360°
- each b subunit goes through all three states
- each b subunit make how many ATP

1

Binding-Change Mechanism for ATP Synthase
The rotation of the γ subunit interconverts the β subunits.
as the γ subunit rotates 360°
- one turn = how many ATP synthesized

3

rotational catalysis

mechanism by which the flow of
protons through Fo causes the c ring to rotate, which causes
the g stalk to rotate, which triggers the subunit conformational
changes in F

n a typical enzyme-catalyzed reaction, reaching which
state between substrate and product is the major energy barrier
to overcome

transition

In the reaction catalyzed by ATP synthase, which is the major energy barrier

release of ATP from the enzyme

Proton Flow Around the c Ring Powers ATP Synthesis
Proton flow occurs through the F0 component of the ATP synthase

Subunit a, next to the c ring, has how many channels that reach halfway
into the a subunit. One half-channel opens to the intermembrane
space and the other to the matrix

2

Protons enter the half-channel facing the proton-rich
intermembrane space, bind to a what residue on one of the
subunits of the c ring, and then leave the c subunit once they
rotate all the way around to face the matrix half channel. Where
they exit moving down the proton gradient

glutamate

Proton-Driven Rotation of the c Ring
the what subunit is
stationary, and the what
ring rotates

a, c

Proton-Driven Rotation of the c Ring

transient protonation
of a key Glu residue in
each what subunit elicits
conformation changes
that drive rotation and
transmit protons

a

each c ring can carry how many protons

1

proton

Proton Flow Around the c Ring Powers ATP Synthesis
The force of the what gradient powers rotation of the c ring

Proton Flow Around the c Ring Powers ATP Synthesis

The rotation of the c ring powers the movement of the γ subunit,
which in turn alters the conformation of the what subunits

β

The rotation of the c ring in vitro
depends on temperature
At 37° C, it rotates how many times/sec

350

protons

Because one complete turn of the F1 subunit produces 3 ATP, the number
of c rings determines the number of what required to synthesize a
molecule of ATP

The c ring of vertebrates consist of how many subunits, making vertebrate ATP
synthase the most efficient known

8

With 8 c ring subunits, our ATP synthase requires 8/3 = 2.67 H+ per ATP
However, remember the cost of one H+ to import phosphate
So, each ATP we make requires the import of how many H+ into the matrix

3.67

ADP

Electrons do not flow through the electron-transport chain unless
what is available to be converted into ATP

acceptor/respiratory control

The regulation of oxidative phosphorylation by ADP is called
what

metabolism

Acceptor control is an example of control of what by
energy charge or mass-action ration

mass-action ratio

[ATP]/([ADP][Pi])

nonshivering thermogenesis

If electron transport is uncoupled from ATP synthesis, heat is generated,
a process called what

uncoupling protein 1 (UCP-1)

Such uncoupling is facilitated in a regulated fashion by what , an integral protein of the inner mitochondrial
membrane, that can let protons across the membrane

Uncoupling occurs in mitochondria in what fat, called what fat
mitochondria

brown

Inhibition of the electron-transport chain

prevents oxidative
phosphorylation by inhibiting the formation of the proton-motive force

Inhibition of ATP synthase

What by inhibiting proton flow prevents electron
transport

Uncouplers

What carry protons across the inner mitochondrial membrane.
The electron-transport chain functions, but ATP synthesis does not
occur because the proton gradient can never form.

Inhibition of the ATP–ADP translocase

What prevents oxidative
phosphorylation